Abstract. Metallodrugs are commonly prescribed to treat cancer, but they have significant off target effects because they kill all quickly dividing cells, including healthy cells. There is a need for new targeted therapies. Light activated ruthenium (Ru) based drugs are promising because they allow for spatial and temporal control of drug activation using the FDA approved technique of photodynamic therapy (PDT). Three Ru based prodrugs have recently entered clinical trials, showing the promise of this approach. New protic Ru complexes have been synthesized and studied which demonstrate light activation and selective toxicity towards breast cancer cells vs. normal cells. Light activation of Ru complexes can generate toxic species by two pathways. 1) Photodissociation of a ligand can generate free ligand and ruthenium with free sites which are both potentially toxic species; this is known as photoactivated chemotherapy (PACT). 2) Alternatively, light driven singlet oxygen generation (via PDT) can lead to oxidative stress and cell death. The aim of this application is to determine the factors that lead to light activated toxicity in new protic Ru photochemotherapy (PCT) agents including whether a PACT vs. PDT process occurs. The long-term goal of this work is to design highly cytotoxic and selective prodrugs that are initially inert but transform into active drugs in the presence of tissue penetrating red light. These prodrugs can target cancer cells due to a combination of enhanced uptake and high levels of oxidative stress present in cancerous cells. Three hypotheses motivate this work. First, protic OH groups on the ligand lead to neutral molecules at physiological pH which can enhance uptake of Ru PCT agents. These OH groups also shift light absorption to longer wavelengths upon deprotonation. Second, the substituent on the ligand determines whether photodissociation vs. singlet oxygen generation occurs upon light activation, which determines the resulting toxicity. Third, breast cancer cells are effectively targeted because of the inherent vulnerability of cancer cells to reactive oxygen species (including singlet oxygen). Three specific aims will probe these hypotheses. 1) Determine how substituent changes control the biological mode of action of Ru PCT agents and their cytotoxicity. 2) Elucidate the mechanism of preferential toxicity towards breast tumor cells. 3) Synthetically design new Ru PCT agents to maximize cytotoxicity and cellular uptake and to red shift light absorption. The proposed research will establish a new generation of Ru PCT agents with selective toxicity towards breast cancer cells and other susceptible cell types. The major innovation of our project is in the rational design of a new class of PCT agents with ideal photophysical, photochemical, biological, and chemical properties. There is potentially a high impact for drug developers beyond the oncology field, in that we are elucidating how protic ligands impact uptake and photodissociation vs. singlet oxygen formation, and this can be used to target other diseases. This project will also be used to train a group of undergraduate and graduate students in biological and chemical collaborative anticancer research.